The present disclosure relates to methods of using a progressive cavity pump as a bioreactor, for example isolating pancreatic islet cells using the progressive cavity pump bioreactor.
A bioreactor is an apparatus used to carry out any kind of bioprocess. For example, a bioreactor can be a vessel in which a chemical process is carried out which involves organisms or biochemically active substances derived from such organisms, such as cell culture, fermentation, tissue growth, or an enzymatic reaction.
A progressive cavity pump is a type of positive displacement pump that transfers fluid by means of the progress, through the pump, of a sequence of small, fixed shape, discrete cavities, as its rotor is turned. This leads to the volumetric flow rate being proportional to the rotation rate and to low levels of shearing being applied to the pumped fluid. An advantage of progressive cavity pumps is their ability to pump solids and liquids simultaneously. Although previously used in a number of industrial applications, progressive cavity pumps have not previously been used as bioreactors for biomedical applications. In particular, progressive cavity pumps have not been used to isolate islet cells from pancreatic tissue.
Diabetes mellitus, often referred to simply as diabetes, is a group of metabolic diseases in which a person has high blood sugar, either because the body does not produce enough insulin, or because cells do not respond to the insulin that is produced.
Chronic diabetes conditions include type 1 diabetes and type 2 diabetes. Type 1 diabetes, also referred to as insulin-dependent diabetes mellitus (IDDM) or juvenile diabetes, results from the body's failure to produce insulin. Type 2 diabetes, also referred to as non-insulin-dependent diabetes mellitus (NIDDM) or adult-onset diabetes, results from insulin resistance, a condition in which cells fail to use insulin properly, sometimes combined with an absolute insulin deficiency. Type 2 diabetes is the most common form.
Diabetes can cause a number of complications affecting the retina, kidney, vascular, gastrointestinal, peripheral, and autonomic nervous systems. The mechanism through which diabetic complications develop is unclear. The generally accepted view for most diabetic complications is that the disease is accompanied by metabolic changes in the affected organ that, in the long term, result in structural alterations.
The islets of the pancreas produce insulin. In type 1 diabetes, the insulin-producing cells in the islets have been destroyed. We do not know how to prevent onset of type 1 diabetes
Treatments for Type 1 Diabetes include insulin therapy, pancreas transplantation, and islet cell transplantation.
Insulin therapy is given by injection or insulin pump. However, even with insulin therapy, many people with type 1 diabetes still have blood glucose levels that are above normal, putting them at risk for the long-term complications of diabetes. Additionally, those who are able to keep their blood glucose levels near normal with insulin therapy often have trouble with low blood glucose (hypoglycemia). After many years, some people lose the early symptoms that warn them that their blood glucose level is dropping (hypoglycemia unawareness) which raises their risk of severe hypoglycemia. Furthermore, some people have what doctors call labile, or brittle, diabetes in which blood glucose levels swing from high to low despite the best insulin plans.
Whole organ pancreas transplant is a major operation and can be associated with complications, such as bleeding, infection, inflammation of the pancreas and clots in the blood vessels around the pancreas. It is most often performed when a patient also needs a kidney transplant. The success rate (long-term insulin independence) with pancreas transplantation was initially low, but increased dramatically in the 1980s. After one year about 85% of pancreas transplant recipients are insulin independent.
Islet transplantation is still in the experimental stages. The advantages of islet transplantation over pancreas transplantation are that it does not require a major operation and the procedure has a small complication rate.
The potential advantage of islet transplantation over administration of insulin by injection is that the transplanted islets would maintain normal blood sugar under all conditions, and would not produce excess insulin resulting in hypoglycemia.
In islet transplantation, islets from a deceased donor are infused (dripped) into a vein in the liver. If the transplant is successful, the islets lodge in the liver and start to produce insulin. The liver is the most common site for islet transplantation, but islets have been transplanted into the peritoneum and spleen as well. Other transplant sites may be possible. Subcutaneous implantation chambers have been proposed as well. The advantage of a subcutaneous implantation chamber is longer viability and retention of islet cells.
In practice, there are problems to overcome in islet transplantation before it can be considered a standard therapy for people with type 1 diabetes.
As with any organ transplant, the recipient of an islet transplant must take drugs every day to keep the body from rejecting the islets. These drugs put the person at risk for infections and certain cancers. They can also cause side effects that range from mild to severe. Some people who received an islet transplant have had to stop taking these medications, because of side effects and then their new islets stopped working.
Sometimes, the transplanted islets never produce insulin. Further, even when the transplanted islets do produce insulin, in the majority of people who receive an islet transplant, the function of the islets deteriorates over time, and they must go back to taking some insulin. Since the number of people who have had successful islet transplants is small, and those have happened within the past decade, it is not clear how long the islets will keep working.
Successful transplants typically require 6,000-9,000 islets per kg bodyweight of the recipient. That is, between 0.5 and 1.0 million islets are needed per transplant recipient. Although a normal human pancreas may have more than one million islets, currently even the most successful isolations seldom yield more than 400,000 islets, often considerably fewer. Therefore, transplant recipients typically require islets from 2-4 donors.
A major obstacle to widespread use of islet transplantation is the shortage of islets. Although organs from about 7,000 deceased donors become available each year in the United States, fewer than half of the donated pancreata are suitable for whole organ pancreas transplantation or for harvesting of islets, enough for only a small percentage of those with type 1 diabetes.
However, researchers are pursuing various approaches to solve the problem of the shortage of islets, including transplanting islets from a single donated pancreas, from a portion of the pancreas of a living donor, or from pigs.
Researchers have transplanted pig islets into other animals, including monkeys, by encapsulating the islets or by using drugs to prevent rejection. Encapsulated porcine islet cells xenotransplanted into at least one human have been shown to continue to produce insulin 10 years after the transplant. Such a product is currently being investigated in Phase II clinical studies in several countries.
The origin and condition of a pancreas, as well as the method of islet cell isolation, can substantially affect the yield and viability of islet cells for either transplant or research purposes. Isolation and purification of islet cells for transplant is presently based on the Ricordi process in which pancreatic tissue is enzymatically digested followed by purification using density gradient separation. (Paget et al., Diabetes Vasc Dis Res 2007, 4:7-12) Currently, although both semi-automated and manual methods are used to isolate pancreatic islet cells from pancreas donors, no fully automated methodology has been developed.
Consequently, there is a need for methods and bioreactors that improve yield and viability of islets isolated from organ donors, especially from human donors.